Traditional methods of laboratory testing of biochemical substances are time-consuming and therefore costly, due to typically repetitive steps required, large numbers of samples that must be tested and retested and the amount and complexity of required laboratory equipment to accomplish the test steps. Many such tests require the sample comprised of particulate or cellular matter suspended in a test medium fluid or in a bodily fluid to undergo active combination with reagents and with intermediate steps of extraction of solid matter and washing with cleansing fluids. The end product of such processes is a residue of solid matter which may be extracted for further analysis by microscopy, reaction in another vessel, radio isotope detection, or spectrophotometery.
Separation of solids from fluid medium may be accomplished by centrifugation, in which a vessel containing the test sample is placed in a mechanical centrifuge and spun until the heavier substances separate to the bottom of the vessel, or simple filtration in which the test fluid is passed through a filtering medium depositing suspended solids which are larger than the filter mesh on the filter, or by gravity separation or absorption in a still laboratory vessel. Filtration methods have the advantage over centrifugation or gravity separation in that they are faster and that several insertions of fluid may be made in the same container and passed through the filter medium, sequentially treating and cleansing the solid material trapped on the filter. In centrifugation or gravity separation the separated fluid must be manually withdrawn from the container before the separated solid may be further washed or treated. Substantial savings of time may be realized therefore by avoiding mechanical removal of fluid from the test container, and in some cases, avoiding removal of the solid material itself to another clean container for further steps. In addition the time required to centrifuge the sample is substantial. The filtration process itself can also be slow however, and may be aided by applying a vacuum to the downstream side of the filter such that the atmospheric pressure differential will force the fluid quickly through the filter. This method can still be time-consuming however, due to limitations of available laboratory equipment and the need for connecting and disconnecting from the laboratory vacuum source the numerous individual containers for samples undergoing tests. Further existing filter manifolds are constructed with exhaust ports whose size permits the fluid to slowly pass through with gravity flow and the fluid will not remain in the container for the length of time necessary to accomplish specific reactions. A separate chamber in such a case would have to be used to accomplish a reaction and transfer to the filter manifold.
Most analytical or quantitative tests of particulate biochemical substances are performed in test tubes or in commercially available containers which provide an array of separate cups drawn together in a grid pattern, usually cast of a plastic, glass or fiberglass solid, enabling simultaneous test of a number of samples. Since the containers are closed wells however, the test steps are subject to the limitations of centrifugation, chemical absorption, or gravity separation and fluids must be drawn off between each test procedure. Efforts to speed this process have involved quite complicated motorized devices to insert and extract fluids from the test wells by passing an array of nozzles over the test wells for inserting fluids and an array of suction ports over the top of the wells to extract fluids.
Finally, all of the methods described above suffer from the dangers of error or contamination of the test samples involved in time-consuming multiple steps, mechanical removal of fluids from the test containers, transfer of the test solids from one container to another, and insertion of instruments such as suction nozzles into the test container.
The device and methology presented herein was developed to facilitate immunodiagnostic tests which were previously difficult to perform. For example, patients with suspected herpes virus infections of the eye usually yield only a few infected cells which can be used to determine the presence or absence of the viral antigens. Because of the numerous manipulations, washing and reaction steps required for immunodiagnostic tests, the cells are often lost in the centrifugation procedures required by these tests. The present invention has been used in conjunction with an indirect radioimmunoassay to detect herpes simplex virus antigens on as few as 195 herpes simplex virus infected human cells. This invention and the accompanying methodology may also be used with direct radioimmunoassays competitive inhibition radioimmunoassays, immunoassays employing fluorescently labeled antibodies, other binding agents (such as staphylococcus aureus protein A) or antigens, or immunoassays employing enzyme labeled antibodies, or other antigens.
Thus it is an object of the present invention to provide a filtration device for chemical and biochemical tests in which a large number of samples may be tested simultaneously.
Another object of the invention is to provide a multiple container test device in which several sequential test steps may be accomplished without removal of the object substance from the test container between test operations.
Another object of the invention is to provide a test method in which large numbers of test samples may be processed without delays for intermediate steps of removal of fluids, transfer of sample containers or mechanical separation.
Another object of the invention is to provide a test method that will separate free reagents from bound reagents rapidly and simultaneously in all samples.
A further object of the invention is to provide an integral vessel for various test steps of different characters, such as filtration and subsequent incubation of samples in the same container to minimize handling of samples in such steps.
Another object of the invention is to provide small reaction chambers in order that only small volumes on the order of 50 microliters of precious reagents need be used.
A final object of the within invention is to provide a simple, inexpensive device which will replace cumbersome, complicated and costly laboratory equipment currently required to perform the tests described above involving numerous sequential test procedures.